EphA2 Receptor Tyrosine Kinase in Malignant Tumor Cell

描述（由申请人提供）：EphA2受体是最近克隆的Eph家族受体酪氨酸激酶（RTKs）的成员，在多种恶性肿瘤中过表达。EphA2受体表达水平与恶性程度一致。EphA2在未转化乳腺上皮细胞系中过表达导致裸鼠肿瘤形成。然而，EphA2在恶性肿瘤中的过表达机制尚不清楚。我们的初步数据显示，在未转化的乳腺上皮细胞MCF10A中，EphA2的过表达会破坏它们在三维培养中形成腺泡样球体的能力。此外，E-cadherin在过表达EphA2受体的MCF10A细胞中表达错位。这些数据表明，高水平的EphA2可能通过破坏细胞-细胞粘附的稳定性来促进恶性行为。反之，在恶性肿瘤细胞中阻断EphA2受体激活可抑制体内RhoA GTPase激活、细胞迁移和肿瘤生长，表明EphA2受体信号是肿瘤恶性发展所必需的。基于这些数据，我们假设EphA2过表达(1)通过改变连接复合物破坏细胞-细胞粘附，(2)通过调节vav3介导的RhoA GTPase活性促进细胞运动，(3)增强肿瘤在体内的侵袭和转移。为了验证这些假设，我们特别提出：(1)确定EphA2过表达破坏细胞-细胞粘附的机制；(2)探讨EphA2过表达细胞的细胞运动性增加是否通过Vav3依赖性激活RhoA GTPase介导；(3)确定EphA2过表达在多期转基因肿瘤模型中促进肿瘤形成和转移是否必要和充分。本项目的成功不仅将为EphA2诱导细胞侵袭转移提供分子基础，还将为EphA2缺乏或过表达对肿瘤进展和转移的影响提供体内临床前数据。

DESCRIPTION (provided by applicant): EphA2 receptor is a member of the recently cloned Eph family receptor tyrosine kinases (RTKs) that overexpressed in a variety of malignant tumors. The level of EphA2 receptor expression consistently correlates with the degree of malignancy. Overexpression of EphA2 in non-transformed breast epithelial line has led to tumor formation in nude mice. However, the mechanism by which EphA2 overexpression in tumor malignancy remains unclear. Our preliminary data show that overexpression of EphA2 in non-transformed breast epithelial cells MCF10A disrupts their ability to form acini-like spheroids in a three-dimentional culture. In addition, E-cadherin expression is mis-localized in MCF10A cells overexpressing EphA2 receptor. These data suggest high levels of EphA2 may promote malignant behavior through destabilization of cell-cell adhesion. Conversely, blocking EphA2 receptor activation in malignant tumor cells inhibits RhoA GTPase activation, cell migration, and tumor growth in vivo, indicating that EphA2 receptor signaling is required for tumor malignancy. Based on these data, we hypothesize that EphA2 overexpression (1) disrupts cell-cell adhesion through modifying junctional complexes, (2) promotes cell motility by modulation of Vav3-mediated RhoA GTPase activity, and (3) enhances tumor invasion and metastasis in vivo. To test these hypotheses, we specifically propose to:(1) identify mechanisms by which EphA2 overexpression disrupts cell-cell adhesion; (2) investigate whether increased cell motility in EphA2 overexpressing cells is mediated by Vav3- dependent activation of RhoA GTPase; and (3) determine if EphA2 overexpression is necessary and sufficient to promote tumor formation and metastasis in multi-stage transgenic tumor models. The success of this project will not only provide molecular basis of EphA2-induced cell invasion and metastasis, but also provide in vivo pre-clinical data on the effect of EphA2-deficiency or EphA2 overexpression on tumor progression and metastasis.